S1PR2 is a direct, repressed target of FOXP1 with proapoptotic activity in DLBCL cell lines. (A-D) S1PR2 expression levels were determined by qRT-PCR (normalized to ACTIN) after 72 hours of FOXP1 depletion in the indicated cell lines. Data are represented as fold change over the negative control siRNA (means + SEM of at least 3 independent experiments are shown). (E-H) ChIP followed by qPCR of 2 FOXP1-bound regions 2.5 kb and 5 kb upstream of the S1PR2 transcription start site that were identified by ChIP sequencing. Data are shown as fold enrichment relative to an unspecific immunoglobulin G control antibody for the 4 indicated cell lines. A locus in the PRR20A gene was used as a negative control.17 Data represent means + SEM of at least 3 independent experiments. (I-K) Viability and apoptosis of the 3 indicated DLBCL cell lines 48 hours posttransfection with an S1PR2 expression plasmid or empty vector. Cell viability was assessed using CellTiter Blue (I) and apoptosis was assessed using Annexin V (J) or cleaved caspase-3 staining (K). Data represent means + SEM of at least 3 independent experiments. (L) A representative enhanced GFP histogram of SU-DHL6 cells after infection with virus particles harboring pIND21-S1PR2. (M-O) S1PR2 expression was induced for 72 hours with doxycycline in SU-DHL6 cells transduced with pIND21-S1PR2 prior to the assessment of S1PR2 transcript levels (M), as well as viability and apoptosis by CellTiter Blue assay and Annexin V staining (N-O); data of 3 independent experiments are shown as means + SEM. *P < .05, **P < .01, and ***P < .001 (2-tailed Student t test). n.s., not significant.
